Studies of the molecular embryology of the lung are likely to provide significant new understanding of the molecular mechanisms of lung development, injury and disease. We have demonstrated that TGF-beta receptor-mediated autocrine/paracrine signaling negatively regulates early lung development. The recent identification of the Smad family signal transducer proteins has unraveled new mechanisms by which TGF-beta signals from the cell membrane to the nucleus, and our preliminary data show that Smad-mediated signaling modulates mouse lung branching morphogenesis and cytodifferentiation. Hypothesis: Specific Smad gene expression regulates TGF-beta signaling, and thus instructs early mouse embryonic lung branching morphogenesis. Specific Aims: Aim 1. To define the developmental and temporo-spatial expression of Smad genes during embryonic lung development (i) in vivo and (ii) in lung explant culture. Aim 2. To determine the molecular mechanism of TGF-beta pathway- restricted Smad2 and Smad3 in regulating embryonic pulmonary branching morphogenesis and cytodifferentiation in serumless culture using both (i) "loss-of-function" and (ii) "gain-of- function" strategies. Aim 3. To define the biological function of the feedback inhibitory Smad6 and Smad7 proteins during embryonic lung branching morphogenesis in culture. (i) Time- and dose-dependent TGF-beta-induced inhibitory Smad6 and Smad7 gene expression patterns will be delineated in lung explant culture. (ii) The antagonistic mechanism by inhibitory Smad6 and Smad7 on TGF-beta signaling during lung development will be determined in embryonic lung culture. Novel molecular mechanisms to control lung morphogenesis and significance to human health: The current proposal will define novel molecular mechanisms in which Smad-mediated signaling regulates embryonic lung branching morphogenesis. The results of this project will provide new rationales for novel therapeutic strategies to modulate TGF-beta signaling during lung development, injury, repair, and disease.